Title: MiR-155 differentially regulates atherosclerosis and metabolically healthy obesity According to the Centers for Disease Control and Prevention, the prevalence of obesity (Ob) has increased to over 35% of the U.S. adult population, or more than 85 million adults (https://www.cdc.gov/obesity/data/adult.html). Ob is an important risk factor for type 2 diabetes mellitus (T2DM), various cancers, nonalcoholic fatty liver disease (NAFLD); cardiovascular disease (CVD), and mortality. In contrast to the classical Ob associated CVD (COB), evidence has shown that some obese individuals are metabolically healthy, termed metabolically healthy obesity (MHO) , and counts for 18-44% of obese adults and 10% adult population. Unlike COB, MHO reports less incidence of atherosclerosis, no insulin resistance (IR) but Ob. MHO is not truly ?healthy? since MHO significantly increase incidences of T2DM, hypertension, and metabolic syndrome (MetS). Therefore, novel therapies are urgently needed to inhibit MHO transition to COB. However, the mechanisms underlying MHO transition to COB remain poorly characterized due to lack of appropriate animal models for MHO transition to COB. The goal of this project is to determine the underlying mechanisms and establish new models of MHO transition to COB. We have long-standing interest in studying metabolic CVD and published many papers on endothelial cell (EC) activation, EC dysfunction, VSMC phenotypic switching, monocyte (MC)/macrophage (M?) recruitment, Foxp3+ Treg in inhibiting CV inflammation, atherosclerosis, Ob/diabetes, chronic kidney disease (CKD), and the roles of caspase-1(Casp1)/inflammmasome in CVD. Therefore, we have sufficient expertise in carrying out this proposal. Our strong preliminary data, new publications showed that: 1) miR155 is significantly upregulated in atherosclerosis aorta and in TNF?-, IL-1?- and lipopolysaccharide (LPS)-stimulated aortic EC; 2) miR155-/-/ApoE-/- mice (DKO) have decreased atherosclerosis, but increase high fat diet (HF)- induced white adipose tissue (WAT) hypertrophy; 3) miR155 KO M? produce less TNF? and IL-1? as others reported; 4) HF significantly increased DKO WAT M? (ATM), proinflammatory adipokines resistin and leptin, which creates systemic proinflammatory conditions and promote MHO transition to COB. These findings suggest that a single cell type, miR155-/- M?, in two different tissue contexts (aorta and WAT), regulates MHO; 5) DKO have no IR and no glucose intolerance, suggesting a MHO status; and 6) we obtained miR155fl/fl mice and are generating cell-specific miR155-/- mice. Our data/publications strongly suggest that DKO have decreased atherosclerosis but have increased HF-induced Ob and ATM without IR, which make DKO as the first model for studying MHO transition to COB. Therefore, the central to be tested is that extended HF feeding transitions MHO to COB by increasing adipose tissue inflammation via increased saturated fatty acids (SFA)-/proinflammatory adipokines - triggered caspase-1 (Casp1) activation in ATM. We will test this hypothesis in three linked aims: Aim 1 To determine whether with 24 week extended HF-fed DKO aortic m? are less inflammatory, but 24 week HF fed DKO WAT and ATM are more inflammatory than ApoE-/- counterparts (relevant studies); Aim 2: To determine whether 24 week extended HF-induced DKO WAT increase SFA, resistin, and leptin, which polarize to M1 ATM by upregulating Casp1/TNF? pathways (mechanistic studies); Aim 3: To determine whether 24 week extended HF-induced DKO mice develop more atherosclerosis than 12 week HF DKO mice; whether HF-induced Casp1-/-/miR155-/-/ApoE-/- triple KO (Casp1 TKO)- and NLRP3-/-/miR155-/-/ApoE-/- (NLRP3 TKO)- WAT produce less resistin52/leptin; whether ATM in two TKO mice produce less IL-1?/TNF?; and whether two TKO mice less Ob and atherosclerotic with extended HF than DKO (verification studies). hypothesis